Successful mucosal pathogens must subvert, invade, or cause damage to the epithelial barrier and then overcome host defense mechanisms, but the strategies by which they accomplish this are incompletely understood. These events are particularly crucial for Pseudomonas aeruginosa (PA), one of the most virulent opportunistic pathogens of humans. New therapeutic modalities are urgently needed as multidrug resistant PA infections become increasingly common, with only a few new PA antibiotics in the pipeline. We have shown that PA subverts host cell polarity in order to overcome the mucosal barrier, but in doing so activates the host innate immune response. Our long-term goal is to understand the underlying mechanism of this process, with the goal of developing new therapies that could be applied to PA and other mucosal pathogens. We have shown that upon binding to the apical surface of polarized epithelial cells, PA forms biofilm-like antibiotic- resistant aggregates. In a ordered and highly regulated process, the host cell responds to the PA aggregate by remodeling the apical surface to form a membranous actin-rich protrusion comprised of basolateral proteins and lipids-resulting in localized inversion of cell polarity. Key determinants of cell polarity are recruited to and required for protrusion formation. Remarkably, these same polarity determinants are also required for activation of the central innate immune effector NF ?B underneath PA aggregates, revealing an unanticipated function for changes in epithelial cell polarity in the activation of the innate immune response. We have recently found that artificially targeting Par3 to the apical membrane, using chemically induced dimerization (CID), is sufficient to recapitulate key aspects of protrusion formation, but is not sufficient to activate NF ?B. Important gaps in our knowledge are: (i) What are the signaling events leading to protrusion formation? (ii) How are cell polarity and innate immunity linked? (iii) What role does the protrusion pathway play in an in vivo system? We propose to Aim 1. Define host molecules that are necessary and sufficient for apical membrane remodeling. Aim 2. Determine how pathogen-evoked changes in cell polarity activate the innate immune system. Aim 3. Elucidate the in vivo correlates of apical membrane remodeling and test potential strategies to interfere with this early response. Relevance. This work is significant because it provides a unique opportunity to elucidate the delicate balance of host-pathogen interactions that lie at the interface between disease establishment and host response. The proposed work will (i) afford the opportunity to develop new therapies against PA infections; (ii) enhance our understanding of how host cell polarity is regulated; and (iii) offer novel insights into how the mucosal barrier distinguishes pathogens from commensals.